PROJECT SUMMARY Chronic kidney disease (CKD) is increasing in the US, now impacting at least one in ten people. CKD typically results in end-stage kidney disease (ESRD), that requires either dialysis or kidney transplantation. Proteinuric glomerular diseases are the major cause of CKD and ESRD and are generally caused by progressive podocyte loss followed by glomerular scarring. Because podocytes are terminally differentiated epithelial cells they cannot regenerate. Therefore, following loss in disease, partial or complete podocyte replacement relies on trans-differentiation of adjacent progenitor cells. These include parietal epithelial cells, and cells of the renin lineage (CoRL), the latter being the focus of this competitive renewal. Unfortunately, endogenous podocyte replacement is often not robust enough to assure protection. Given this situation, and to address the unmet need to augment podocyte replacement through discovery of new targets, our long-term goal is to understand the mechanisms underlying podocyte regeneration, and identify therapeutic options towards restoring podocyte numbers and preventing glomerular scaring. We recently reported that CoRL are pluri-potent progenitors that can trans-differentiate towards podocytes, parietal epithelial cells, mesengial cells and pericytes. We have also shown that the transcription factor Wt1 is a critical player and is necessary for CoRL proliferation and migration, and its trans-differentiation to a podocyte fate. However, the precise molecular mechanism(s) underlying this CoRL biology are still poorly understood. To resolve this, the overall objective of this application is to investigate the mechanisms and pathways to augment CoRL progenitor function and its trans- differentiation into podocytes. Our central hypothesis is that we can ramp up the ability of these CoRL to become podocytes, and replace those lost by modulating distinct signaling systems such as Wnt signaling. To test this, we will pursue two Specific Aims: (1) Test the hypothesis that the progenitor capacity of cells of renin lineage (CoRL) to trans-differentiate to a podocyte fate is governed by the balance of WT1 and Wnt signaling, and (2) Identify pathways underlying the pluripotency of cells of renin lineage towards different adult kidney cell type fates in glomerular diseases. The approach is innovative as it is based on a unique ability to culture CoRL ex vivo, innovative bioengineered cell culture devices, a newly developed dual-reporter mice to definitively trace CoRL-podocyte trans-differentiation, and Quality-by-Design (QbD)-based methodologies to identify complex and robust culture conditions. The research is significant as it provides an in depth understanding of the potential of CoRL to differentiate into multiple kidney cell types even in adults. In addition, it will advance the field by identifying potentially novel therapeutic options towards restoring podocyte numbers, and in doing so, preventing and even reversing glomerular scaring in proteinuric glomerular diseases.